Protein synthesis inhibitors exhibit a nonspecific effect on phenobarbital-inducible cytochome P450 gene expression in primary rat hepatocytes.
Previous investigations have indicated that de novo protein synthesis is a critical requirement for phenobarbital (PB) induction. We reexamined this issue in PB-responsive primary rat hepatocyte cultures using a broader array of protein synthesis inhibitors and experimental end points. Anisomycin, cycloheximide, emetine, puromycin, and puromycin aminonucleoside, a negative analog, were evaluated for their respective effects on protein synthesis and the PB-induction process. All of the inhibitors effectively repressed de novo protein synthesis in the cells in a concentration-dependent manner. However, anisomycin only minimally effected PB induction, ascertained though the measure of CYP2B1, CYP2B2, and CYP3A1 mRNA levels. The inactive agent, puromycin aminonucleoside, produced marked repression of the PB-induction response. Results from further experiments demonstrated that these protein synthesis inhibitors stimulated rapid and differential phosphorylation of the stress-activated protein kinase/c-Jun kinase (SAPK/JNK) pathway, indicating nonselective actions on cellular processes. Puromycin aminonucleoside was without effect on these pathways, despite its efficacy as an inhibitor of PB induction. These results demonstrate that de novo protein synthesis is not a requirement for PB induction, nor is activation of the SAPK/JNK kinase cascade responsible for down-regulating PB responsiveness in primary hepatocytes.
Sidhu
J S
JS
Department of Environmental Health, University of Washington, Seattle, Washington 98195, USA.
Omiecinski
C J
CJ
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002478:Cells, Cultured; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics*; D004789:Enzyme Activation; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D048031:JNK Mitogen-Activated Protein Kinases; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D020928:Mitogen-Activated Protein Kinases; D010634:Phenobarbital / Q000494:pharmacology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D051381:Rats; D015398:Signal Transduction
D011500:Protein Synthesis Inhibitors; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C465138:Cyp3a23-3a1 protein, rat; D051544:Cytochrome P-450 CYP3A; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D010634:Phenobarbital